This invention relates to methods for detecting polypeptides in gel matrices.
Polyacrylamide gel electrophoresis is one of the major methods for separating polypeptides based on their relative size and overall charge. After separation, the location of the polypeptides in the gel is determined by staining the polypeptides; alternatively, the polypeptides may be prelabelled, for example, by radioisotopes, and their location determined by autoradiography.
One of the most common polypeptide stains is Coomassie blue. Another stain is silver, for example as described by Morrissey (U.S. Pat. No. 4,468,466). Silver staining is about one hundred-fold more sensitive than Coomassie blue staining. Hager et al. (1980, Anal. Bioc. 109:76) disclose staining gels using 0.25 M KCl and 1 mM DTT (dithiothreitol).
It is also possible to stain the polyacrylamide gel rather than the polypeptides, so that the polypeptide bands appear as transparent (or clear) regions within a white (or opaque) gel. For example, Higgens et al. (1978, Anal. Biol. 93:257) disclose staining polyacrylamide gels containing 0.1% SDS (sodium dodecyl sulfate) with 4M sodium acetate. In place of sodium acetate, Higgins discloses use of NaCl, KCl, NH.sub.4 Cl, (NH.sub.4).sub.2 SO.sub.4 and potassium acetate.
Casero et al. (1985, Electrophoresis 6:367) disclose the use of colloidal gold as a negative stain.
Barg (U.S. Pat. No. 3,892,841) describes the use of soluble metal salts of zinc or copper to enhance the visibility of immunodiffusion precipitin bands. Precipitated protein is positively stained with CuCl.sub.2 in the presence of Tris buffer between pH5.5 and 7.5.